Axonal regulation of myelination by neuregulin 1

Klaus-Armin Nave1,2 and James L Salzer3

Neuregulins comprise a family of epidermal growth factor-like ligands that interact with ErbB receptor tyrosine kinases to control many aspects of neural development. One of the most dramatic effects of neuregulin-1 is on glial cell differentiation. The membrane-bound neuregulin-1 type III isoform is an axonal ligand for glial ErbB receptors that regulates the early Schwann cell lineage, including the generation of precursors. Recent studies have shown that the amount of neuregulin-1 type III expressed on axons also dictates the glial phenotype, with a threshold level triggering Schwann cell myelination. Remarkably, neuregulin-1 type III also regulates Schwann cell membrane growth to adjust myelin sheath thickness to match axon caliber precisely. Whether this signaling system operates in central nervous system myelination remains an open question of major importance for human demyelinating diseases.
Addresses 1 Max Planck Institute of Experimental Medicine, D-37075 Goettingen, Germany 2 Hertie Institute of Multiple Sclerosis Research, Goettingen, Germany 3 Departments of Cell Biology and Neurology, and the Molecular Neurobiology Program, Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, NY 10016, USA Corresponding author: Nave, Klaus-Armin (nave@em.mpg.de)

In this review, we describe recent progress in elucidating the mechanisms by which motor and sensory axons in the peripheral nervous system (PNS) regulate the development and differentiation of Schwann cells, most strikingly during myelination. Unexpectedly, a single growth factor, neuregulin-1 (NRG1), has emerged as the pivotal signal that controls Schwann cells at every stage of the lineage.

Neuregulin-1 and ErbB receptors

The Neuregulin-1 (NRG1) family comprises more than 15 membrane-associated and secreted proteins [4,5]. These are derived from one of the largest mammalian genes (on human chromosome 8p22 and mouse chromosome 8A3) and are generated by use of multiple transcription sites and by extensive alternative RNA splicing [6]. All NRG1 isoforms share an epidermal growth factor (EGF)-like signaling domain that is necessary and sufcient for activation of their receptors. NRG1 isoforms are subdivided into several subtypes on the basis of their distinct amino-termini [4,7]. NRG1 type I (also known as heregulin, neu differentiation factor, or acetylcholine receptor-inducing activity [ARIA]) and NRG1 type II (also known as glial growth factor [GGF]) have N-terminal immunoglobulin-like domains. Transmembrane forms of NRG1 undergo proteolytic cleavage by metalloproteinases (MP), including TACE (tumor-necrosis factor-a-converting enzyme) [8]. As a consequence, NRG1 type I and II are shed from the neuronal cell surface and function as paracrine signaling molecules (schematically depicted in Figure 1). NRG1 type III is dened by its cysteine-rich domain (CRD), which functions as a second transmembrane domain. Consequently, NRG1 type III remains tethered to the cell surface after cleavage and functions as a juxtacrine signal [9]. In addition, exons encoding shorter amino termini of NRG1 have been identied by sequence analysis (referred to as types IVVI), but these isofoms have not been further characterized [10]. NRG1 expression is not specic to the nervous system, it also has a major role in cardiac and mammary tissue development (reviewed in [11]). Indeed, mice lacking NRG1, or its receptors (ErbB2, ErbB3 and ErbB4), are embryonic lethal because NRG1ErbB signaling is essential for cardiac development. Within the nervous system, NRG1 types I and III are the most abundant forms and have been detected in many projection neurons, most notably in spinal motor neurons and dorsal root ganglia (DRG) neurons, but also in glia [4,12]. In addition to the axonglia signaling detailed below, the proposed functions of NRG1 include the development of motor endplates, migration of interneurons, and
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Current Opinion in Neurobiology 2006, 16:492500 This review comes from a themed issue on Neuronal and glial cell biology Edited by Kelsey C Martin and Elior Peles Available online 7th September 2006 0959-4388/$ see front matter # 2006 Elsevier Ltd. All rights reserved. DOI 10.1016/j.conb.2006.08.008

Introduction
Reciprocal interactions between neurons and glia are crucial for the organization and function of the nervous system, from neurogenesis in embryonic development to synaptic plasticity in the adult brain. Glial cells that synthesize myelin are essential for normal motor and cognitive functions, with the ne tuning of myelination contributing to the millisecond precision of the nervous system [1]. Furthermore, myelin-forming glial cells are also required for the long-term integrity of axons, independently of myelin itself [2,3]. Axons, in turn, crucially regulate the behavior of myelinating glia: that is, Schwann cells and oligodendrocytes. However, the molecular mechanisms by which neurons and glial cells communicate remain poorly understood.
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Axonal regulation of myelination by neuregulin 1 Nave and Salzer 493

Figure 1

NRG1 isoforms: membrane disposition and signalling. (a) Types I and II are synthesized as single pass transmembrane proteins; Type III has two transmembrane domains. (b) With metalloproteinase (MP) cleavage, types I and II are shed as paracrine signals, type III remains tethered through its cysteine rich domain (CRD) and is a juxtacrine signal; this cleavage is enhanced by neurotrophins released by Schwann cells. The cytoplasmic domain undergoes further cleavage stimulated by binding of ErbB receptors to NRG1, followed by translocation to the nucleus. See [4] for additional details.

synaptogenesis and synaptic plasticity in the CNS [5]. Many NRG1-expressing neurons also express transcripts for NRG2 and NRG3, two structurally related growth factors with EGF-like signaling domains, the function of which in the nervous system remains largely unknown [12,13].
ErbB receptors

adaptor molecules and activation of downstream signaling pathways (see below). Whereas Schwann cells principally express ErbB2 and ErbB3, cells in the oligodendrocyte lineage express all three ErbB receptors, in a developmentally regulated manner, in addition to the EGFR (ErbB1), indicating signicant complexity of potential ErbB receptor heterodimers and downstream signaling events in these cells [16,17].
NRG1 back signaling

NRG1 isoforms mediate their effects by binding to ErbB receptors, members of the EGF receptor superfamily [14]. NRG1 binds to either ErbB3, which lacks an active kinase domain, or ErbB4, which has such a kinase domain; each receptor, in turn, can heterodimerize with ErbB2, which cannot bind NRG1 directly but also has an active kinase domain. ErbB receptors dimerize not by virtue of a bridging effect of NRG1, but following a ligand-activated conformational change in the ectodomain of ErbB3 or ErbB4 [15]. Crystallographic data indicate that ErbB2 constitutively exposes a dimerization loop required to form heterodimers with ligand-activated ErbB3 or ErbB4 receptors. Because the ability of ErbB2 to form homodimers is poor, and ErbB4 is minimally expressed by Schwann cells, ErbB2ErbB3 is the relevant NRG1 Schwann cell receptor. NRG1 binding induces ErbB2 ErbB3 heterodimer formation, which leads to receptor cross-phosphorylation, recruitment of SH3-containing
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Intriguingly, NRG1 might also signal bidirectionally. Binding of recombinantly produced soluble ErbB receptors to membrane-bound NRG1 type III of transfected neurons results in proteolytic cleavage of NRG1, releasing its cytoplasmic C-terminal domain (CTD) from the membrane [18]. Remarkably, the CTD then rapidly translocates into the cell nucleus of the cultured neurons where it activates transcription and enhances survival. Whether NRG1 backsignaling occurs in vivo, particularly when NRG1 expressed on the axon is engaged by ErbB receptors of myelinating glia, remains to be established. It will also be important to determine whether loss of the anti-apoptotic effect of the CTD is normally responsible for the degeneration of dorsal root ganglion (DRG) and motor neurons that is observed in NRG1 null mutant mice. Alternatively, conventional NRG1 (forward) signaling could elicit
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reciprocal trophic support of ensheathed axons, such as through NRG1-stimulated release of neurotrophins by glia [19]. Finally, the cytoplasmic tail of NRG1 has been reported to interact directly with LIM kinase, a regulator of the actin cytoskeleton [20]; the biological signicance of this interaction is not yet established.

indicate that nal matching of SCPs to axons is mediated through competition for a NRG1-mediated survival signal [2426]. NRG1 type III is the key isoform required for SCP survival and migration during early embryogenesis [26].
Threshold levels of NRG1 type III are an instructive signal for myelination

The role of NRG1 in Schwann cell myelination

NRG1 has a crucial role at essentially every developmental stage of Schwann cells, as rst indicated by both culture studies and analysis of knockout mice [21,22]. These functions include promoting the gliogenic fate of trunk neural crest cells, the migration of Schwann cell precursors (SCP) along axons, and their subsequent proliferation and survival induced by axons. A recent study, analyzing zebrash ErbB mutants, strongly supports the key role of NRG1ErbB signaling in SCP proliferation and directed migration along axons although, surprisingly, not for survival [23]. This result contrasts with studies in rodents and chick, which
Figure 2

Once generated, SCPs differentiate into mature Schwann cells that either ensheath multiple small, unmyelinated axons, forming a Remak bundle, or sort larger axons into a 1:1 relationship that they subsequently myelinate (schematically summarized in Figure 2). These alternative phenotypes are distinguishable not only by their anatomic relationship to the axon but by the repertoire of transcription factors and proteins that Schwann cells express. The axon determines this binary choice in Schwann cell phenotypes, as rst demonstrated in classic studies performed over a century ago in which myelinated and nonmyelinated nerves were cross-anastomosed [27]. As

Axonal NRG1 regulates successive steps of Schwann cell differentiation. (a) Schwann cells (in blue) arise from neural crest precursor cells (in green) and interact with both large and small caliber axons of spinal motor and sensory neurons. During embryogenesis, NRG1 on the axon regulates Schwann cell development by activating ErbB signaling cascades, thereby promoting Schwann cell differentiation and expansion. The amount of NRG1 type III on the axon detected by committed Schwann cells, which is a function of axon size and NRG1 levels, then drives them either into segregating single axons and myelination (top), or into a non-myelinating phenotype and formation of a Remak bundle (bottom). Above threshold levels, NRG1 type III signals axon size to Schwann cells to optimize myelin sheath thickness. (b) In mouse mutants lacking NRG1 (/), in heterozygous NRG1 (+/) mice, and in transgenic NRG1 overexpressing mice, the amount of myelin made by Schwann cells varies directly as a function of axonal NRG1 type III levels (indicated by yellow dots) rather than as a function of axon diameter. See [30,32] for further details. Current Opinion in Neurobiology 2006, 16:492500 www.sciencedirect.com

Axonal regulation of myelination by neuregulin 1 Nave and Salzer 495

myelination in the PNS typically commences around axons with a diameter of 1 mm or greater, it was initially posited that a critical axonal diameter was the trigger for Schwann cell myelination [28,29]. Recent studies by Taveggia et al. [30] on the role of NRG1 in myelination suggest that the level of NRG1 type III on the axon, rather than axon diameter per se, is the key instructive signal for myelination. Expression of NRG1 type III on the axon correlates with the ensheathment fate of axons: unmyelinated, autonomic neurons express low levels of NRG1 type III on the axon surface, whereas brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) dependent dorsal root ganglion (DRG) neurons, the axons of which are heavily myelinated, express high levels. DRG axons from NRG1 type III null mice are not myelinated by Schwann cells in cocultures and do not induce myelinspecic structural proteins or transcription factors, thus demonstrating that axonal NRG1 type III is essential for myelination [30]. Furthermore, mice haploinsufcient for NRG1 type III have a signicantly higher proportion of axons that are persistently unmyelinated [30]. Accordingly, pharmacologic inhibitors of ErbB receptors demonstrated a requirement for NRG1 signaling for the initiation of myelination in zebrash [23]. Strikingly, forced expression of NRG1 type III in the post-ganglionic bers of sympathetic neurons converts these normally unmyelinated bers to myelinated ones in vitro [30]. Likewise, in transgenic mice, overexpression of NRG1 type III induces earlier onset of myelination in the PNS than normal, and results in myelination of very small-caliber C-ber axons that would normally be unmyelinated (M Schwab and KA Nave, unpublished). Taken together, these results suggest that threshold levels of NRG1 type III provide the long sought instructive signal that triggers Schwann cell myelination (Figure 2).
NRG1 type III levels regulate myelin thickness

axon surface. Altering this communication impacts on the g-ratio of myelinated axons. Thus, heterozygous NRG1 type III null mutant mice, that display roughly 50% of the NRG1, are hypomyelinated, have commensurate reduction in myelin transcription factors and exhibit reduced nerve conduction velocities [30,32]. By contrast, transgenic mice that overexpress NRG1 in dorsal root ganglia and motor neurons (under control of the neuronal ThyI promoter) become hypermyelinated [32]. Axon-toSchwann cell signaling appears specic to NRG1 type III, as transgenic mice that overexpress the secreted NRG1 type I are not hypermyelinated; this specicity might reect a requirement for juxtacrine signaling characteristic of the type III isoform [9,30,32]. Whereas NRG1 has emerged as the rate-limiting factor of myelin growth control, ErbB2 and ErbB3 are expressed at saturating levels [32]. They must be dramatically reduced to disrupt myelination, such as in a conditional mouse mutant carrying the Schwann cell-specic null mutation of the ErbB2 gene [33]. Similarly, mice that express a dominant-negative ErbB receptor in Schwann cells under control of the 20 ,30 -cyclic nucleotide phosphodiesterase (CNPase) promoter are hypomyelinated and have thinner myelin sheaths than normal [34]. The length of myelin internodes in mice expressing a dominant negative ErbB receptor is also shorter than normal [34], in contrast to mice with reduced NRG1 dosages that have internodes of normal length [32]. Taken together, these studies support a model in which threshold levels of NRG1 type III are required to trigger myelination. Above this threshold, the amount of myelin formed is graded to the amount of NRG1 type III presented by the axon to the Schwann cell an amount likely to reect both the concentration of NRG1 type III at the membrane and the axon surface area (a function of axon diameter). These results also provide a mechanism by which axons, through differing levels of NRG1 type III, coordinate Schwann cell numbers to their alternative phenotypes. Thus, axons that express higher levels of NRG1 type III generate the additional Schwann cells required to establish the 1:1 relationship characteristic of myelinated bers and, subsequently, drive axon segregation and myelination.
Role of NRG1 in myelin maintenance and the injury response

NRG1ErbB signaling has also been recruited for the important regulatory step associated with the nal stage of Schwann cell differentiation: the quantitative control of myelin sheath thickness. When aiming for the most rapid impulse propagation, which is a function of axon caliber and myelination, the optimal myelin thickness is reached when the g-ratio (i.e. the numeric ratio between the diameter of the axon cylinder and that of the myelinated axon) is close to 0.68. For most vertebrates, this ratio is remarkably well maintained for peripheral axons and their myelinating glia independent of the specic axon diameter as rst noted by Donaldson and Hoke [31]. This requires axon size to be perceived by myelinating Schwann cells, to make the correct number of myelin wraps. Michailov et al. [32] demonstrated that this axon size information is encoded, at least in part, by the amount of membrane-associated NRG1 type III displayed on the
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Recent studies have addressed whether ongoing NRG1 signaling is required to maintain the axon-myelinating Schwann cell unit after it has formed, and whether NRG1 has a role during Wallerian degeneration. To address the rst question, Atanasoski et al. [35] ablated the ErbB2 receptor gene in adult myelinating Schwann cells using tamoxifen-inducible Cre recombinase, expressed under the control of the proteolipid protein promoter (PLP); they found, surprisingly, that myelin
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sheaths were unaffected even after two months. Results from this study strongly suggest that ongoing ErbB2 signaling, and by inference NRG1 activity, is dispensable for maintenance of the myelin sheath in the adult. This result is also consistent with the maintenance of myelin sheaths in neuronSchwann cell cocultures even when continuously treated with pharmacologic inhibitors of signaling pathways activated by NRG1 [36]. NRG1 has also been considered a candidate to mediate the Schwann cell response to injury as it is persistently expressed by adult axons [30,32,37], and could therefore be released during Wallerian degeneration to stimulate Schwann cell proliferation. In support, ErbB2 is rapidly activated (phosphorylated) within minutes of injury [38] followed by a delayed, and sustained (over days), upregulation of ErbB receptors and NRG1 isoforms by Schwann cells [37,39]. In addition, several investigators have reported that addition of NRG1 type II results in demyelination and proliferation in cocultures [38,40,41] or when expressed as a transgene in Schwann cells under the control of the P0 promoter in vivo [42]. These ndings suggest that NRG1 isoforms could function as an injury signal, or contribute to nerve pathology, when either misexpressed or aberrantly presented to the outer (abaxonal) Schwann cell membrane. Important support for a role of NRG1ErbB signaling in the injury response was also provided by studies in which pharmacologic inhibition of ErbB2 receptors blocked Schwann cell proliferation after injury in vitro and in vivo [38]. Surprisingly, in a second study in which ErbB2 expression was conditionally ablated in vivo before injury, no effect on Schwann cell proliferation during Wallerian degeneration was seen [35]. The reasons for the discrepancy between these two studies is not yet clear.
NRG1 regulates axonSchwann cell interactions in Remak bers

of impaired glial-derived neurotrophic factor (GDNF) expression by Schwann cells [43]. These observations also reveal that ensheathing glial cells in the PNS, as in the CNS [3], have a primary axon-protective function, independent of myelination.
NRG1-activated signaling pathways and myelination

The signaling pathways by which axons, through NRG1, promote Schwann cell myelination have also begun to emerge. NRG1 robustly activates mitogen-activated protein (MAP) kinase and PtdIns 3-kinase pathways in cultured Schwann cells (reviewed in [5,44]). Axonal contact also robustly activates these signaling pathways in Schwann cells [36] NRG1 type III is the key neuronal signal that mediates PtdIns 3-kinase activation, whereas other signals, distinct from NRG1 type III but not yet identied, activate MAP kinase [30]. Activation of the PtdIns 3-kinase pathway, and its downstream effectors, notably the serine-threonine kinase Akt, are crucial for the trophic, proliferative and differentiative responses of Schwann cells to axons and NRG1. Pharmacologic inhibition of PtdIns 3-kinase blocks the ability of axons to promote Schwann cell proliferation, survival and induction of myelination [36]. Expression of dominant negative forms of PtdIns 3-kinase or Akt in Schwann cells inhibits myelination in vitro, whereas overexpression of PtdIns 3-kinase or activated Akt promote myelin protein expression in vitro and enhanced myelin sheath formation during regeneration in vivo [45]. In contrast to its promyelinating effects, activation of MAP kinase pathways inhibits myelination [45] and causes myelinating Schwann cells to dedifferentiate and proliferate [41]. This has led to the notion that the balance between these two major signaling pathways determines the differentiative state of Schwann cells [45]. Two important signals that also regulate Schwann cell differentiation, laminin in the extracellular matrix (ECM) and cAMP-dependent pathways, were recently implicated in NRG1-dependent signaling. Mice decient in laminin expression fail to ensheath axons appropriately and exhibit markedly reduced ErbBPtdIns-3 kinase signaling associated with increased Schwann cell apoptosis [46]. Impaired ErbB signaling might result from aberrant physical interactions between Schwann cells and axons and/or altered co-signaling by ECM components. Laminin, through interaction with integrin receptors, has been reported to switch the signaling pathways mediated by soluble NRG1 isoforms in cultured oligodendrocytes [47] and so might also qualitiatively affect Schwann cell signaling activated by NRG1. Moreover, addition of cAMP analogs to cultured Schwann cells mimics the effect of axonal contact and has synergistic effects with NRG1 [22]. Elevated cAMP increases ErbB receptor expression and leads to sustained activation of NRG1-dependent signaling pathways [48,49].
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Although axonal NRG1 is the primary signal for immature Schwann cells to adopt the myelin-forming phenotype ([30] and Schwab MH, Humml C, Nave K-A, unpublished), NRG1ErbB signaling is also important for the organization and function of adult non-myelinating Schwann cells. Adult heterozygous NRG1 Type III null mice exhibit impaired sorting of unmyelinated axons into separate Schwann cell pockets, which instead frequently persist as large bundles [30]. These results indicate a more general role for NRG1 signals in axon sorting by Schwann cells. Overexpression of a truncated (dominantnegative) ErbB4 receptor under control of the glial brillary acidic protein (GFAP) promoter in Remak Schwann cells also leads to signicant abnormalities. These include progressive loss of small C-ber axons, which leads to death of DRG neurons, pain insensitivity and a novel neuropathy phenotype; neuron cell loss might reect signicant reductions of glial support, potentially because
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A role for neurotrophins in axonal NRG1 signaling

Neurotrophins exert multiple effects on developing glia, including Schwann cells and oligodendrocytes. A remarkable ability to stimulate Schwann cell differentiation in vivo was observed by Grifn and co-workers [50], who found that the injection of glial-derived growth factor (GDNF) into rats caused non-myelinating Schwann cells to proliferate and even to myelinate some of the very small caliber C-ber axons. These experiments do not distinguish between a direct effect on glia and an indirect one (e.g. by stimulating axons to express NRG1). Esper and Loeb [51] demonstrated that nerve growth factor (NGF) and GDNF, both of which are expressed by Schwann cells, induce the rapid release of NRG1 from the axons of cultured DRG neurons and motor neurons. Although the molecular mechanisms are not fully understood, this neurotrophin-inducible NRG1 release occurs within minutes, is dose-dependent, and can be mimicked by protein kinase C activation. It probably involves regulated proteolytic processing of cleavable NRG1 isoforms, such as transmembrane type I and II isoforms. Taken together with evidence that NRG1 promotes GDNF expression by Schwann cells in Remak bers [43], it appears that a regulatory loop of glial-axon-glial signaling through growth factors determines both neuronal survival and Schwann cell differentiation in the peripheral nervous system. Independently, Chan et al. [52] showed that neurotrophins have strong but opposing effects on myelination by Schwann cells versus oligodendrocytes. In DRG Schwann cell cocultures, adding NGF to the medium promoted myelination, whereas in DRGoligodendrocyte cocultures, NGF had an inhibitory effect. The requirement of (neuronal) TrkA receptors and the results of experiments with Campenot chambers suggest that neurons, not glial cells, are the crucial target cells of NGF signaling. Although the idea that neurons are stimulated by NGF is in agreement with the data of Esper and Loeb [51], the divergent responses of Schwann cells and oligodendrocytes to DRG neurons treated with NGF is unexpected. It contradicts the view that axons that project from the CNS into the PNS (and vice versa) are uniformly myelinated because they provide the same signals to oligodendrocytes and Schwann cells. Modifying this view might help to elucidate temporal differences in the onset of CNS and PNS myelination. It is not yet known whether some of these effects involve NRG1 signaling. The relevance of these in vitro observations in normal development remains to be determined, possibly by careful analysis of existing mouse mutants.

issue is yet to be resolved. However, several studies suggest that NRG1ErbB signaling might regulate oligodendrocyte development and provide insights into its potential role during differentiation. Initial studies in which oligodendrocyte progenitor cell (OPC) cultures were supplemented with soluble NRG1 isoforms suggested that NRG1 has trophic and mitogenic effects on cells in the oligodendrocyte lineage; their effects on differentiation varied considerably between studies (reviewed in [53]). Analysis of the role of NRG1 in a more physiologic context has been confounded by the early embryonic lethality of NRG1 and ErbB receptor knockouts. As an alterative strategy, the role of NRG1 and ErbB receptors in oligodendrocyte development has been analyzed during the ex vivo development of embryonic spinal cord from mice decient in these proteins. These studies suggest a complex role of NRGErbB signaling in the oligodendrocyte lineage. OPC development was reported to be markedly decient in spinal cord explants from NRG1 null mice [54]. Loss of ErbB2 likewise impaired OPC differentiation [55]. However, loss of ErbB3 had no effect on oligodendrocyte differentiation or myelination [56], whereas loss of ErbB4 paradoxically enhanced oligodendrocyte differentiation [17]. It is probably the case that altered rather than completely absent ErbB signaling in these receptor knockouts accounts for these widely divergent effects on oligodendrocyte differentiation. Mice that express a chimeric, dominant negative ErbB receptor protein to inhibit ErbB signaling broadly in the oligodendrocyte lineage had increased numbers of progenitor cells, reduced numbers of mature oligodendrocytes, and were hypomyelinated [57]. This hypomyelination suggests that ErbB signaling, and potentially NRG1 or related ligands, could regulate CNS myelination. In the future, analyses of conditional NRG1 and ErbB receptor knockouts are expected to more precisely delineate the important issue of the role of this signaling pathway in the oligodendrocyte lineage.

Clinical implications
Null mutations of the NRG1 gene and its receptors are embryonically lethal in mice [7], suggesting that human NRG1 loss-of-functions are unlikely to be a primary cause of disease. However, the many roles of NRG1 in glial cell development suggest that dysregulated NRG1 expression (or abnormal Nrg1-ErbB-PI3K signaling) contributes to disorders of myelin as a disease modier or a genetic risk factor. Although this is an interesting possibility, it remains speculative as no evidence directly links NRG1 to myelin disorders as of yet. One potential example is the inherited demyelinating neuropathies (Charcot-Marie-Tooth [CMT] disease type 1) of the peripheral nervous system that result from the abnormal expression of myelin membrane
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The role of NRG1 in oligodendrocyte development

An obvious question is whether NRG1 type III also regulates oligodendrocyte myelination this important
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proteins [58,59] including P0, PMP22, and connexin32. Non-specic pathological features of CMT1 include dysmyelination and onion bulb formation by supernumerary Schwann cells. It is intriguing to speculate that perturbed physical interactions, for example between axons and ensheathing glial membranes, secondary to abnormal expression of myelin proteins or gap junction proteins, alter NRG1-ErbB-PI3K signaling, and contribute to the CMT1 pathology. In the adult central nervous system, multiple sclerosis (MS) lesions often fail to remyelinate, despite the presence of oligodendrocytes and OPC [60]. If remyelination occurs at all, the resulting sheaths are abnormally thin [61]. This is reminiscent of the thin sheaths of remyelinated peripheral nerves [62] and the hypomyelination seen in the PNS of mice with reduced NRG1 gene dosage [30,32]. An intriguing possibility is that the steady-state levels of NRG1 or related growth factors on myelinated axons in the adult CNS (or PNS) are reduced below the level required for robust remyelination, particularly in demyelinated regions [63,64]. These ideas can be experimentally tested in mice with a genetically modied NRG1 expression level, when challenged by either an experimental autoimmune encephalomyelitis (EAE) or a toxininduced demyelination to undergo spontaneous remyelination. Earlier reports that the systemic application of recombinant NRG1 improved the course of EAE in mice, with respect to disease onset and severity [63,65], contrasts with the inability to improve the degree of remyelination in toxin-induced rat CNS lesions [66]. This suggests that the benecial effects of systemic NRG1 administration have been indirect, for example, through immune modulation. It is our experience from experiments in the peripheral nervous system that functional NRG1 signaling to myelinating glia must be presented directly by the axon [30,32], whereas ectopic stimulation by soluble NRG1 might even have detrimental effects on myelination [40]. This might reect the requirement for NRG1 to signal in a juxtacrine mode, or that NRG1 only promotes oligodendrocyte differentiation in the context of lamininintegrin signaling at the axonglial interface [47]. Finally, there are intriguing links between NRG1 expression, myelination and neuropsychiatric disorders. Stefansson and coworkers [67] reported the association of single nucleotide polymorphisms (SNPs) in the 50 region of the NRG1 gene with schizophrenia in the Icelandic population, an association now conrmed in other populations [68]. The molecular consequences of the crucial NRG1 at risk haplotype are still unclear as the SNPs all correspond to non-coding variants, but recent evidence suggests alterations of expression of NRG1 types I, III and IV [6,69]. Because NRG1ErbB signaling in the CNS affects oligodendrocyte development in addition to neuroblast migration and glutamatergic synapse function, NRG1 is a
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plausible susceptibility gene to contribute to human schizophrenia [70]. Interestingly, CNS myelin abnormalities have also been independently associated with schizophrenia and other neuropsychiatric diseases on the basis of gene array and imaging studies [71,72], adding credence to a potential glial contribution.

Outlook and conclusions

The identication of NRG1 as the axonal signal that drives the entire Schwann cell lineage, including myelination, is an important milestone that will facilitate elucidation of the mechanisms that underlie the morphogenetic and transcriptional events of myelination. In the PNS, important remaining questions include how NRG1-dependent activation of PtdIns 3-kinase initially promotes proliferation but later drives differentiation of Schwann cells, how NRG1 signaling strength regulates the binary choice of Schwann cell phenotypes, what limits NRG1 signaling once myelination is complete, and whether perturbations of NRG1 signaling contribute to neuropathology. Future studies should also clarify the role of NRG1, and related genes, in CNS myelination, including the question of whether these axonal factors limit the efcacy of remyelination in the adult brain. Finally, the temporal and quantitative control of NRG1 expression by neurons remains to be explored. Elucidating these questions promises to provide important insights into the role of axon glial signaling in demyelinating disease and myelin repair.

Acknowledgements
Owing to space limitations, we regret any omissions in citing other relevant publications. We thank C Birchmeier, D Falls, C Lai, J Loeb, M Schwab, and C Taveggia for insightful discussions and for comments on the manuscript. Work from the authors laboratories cited in this review has been supported by grants from the Duetsche Forschungsgemeinschaft (Center for the Molecular Physiology of the Brain), National Institutes of Health, and the National Multiple Sclerosis Society.

References and recommended reading

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